Document Detail


A novel micropit device integrates automated cell positioning by dielectrophoresis and nuclear transfer by electrofusion.
MedLine Citation:
PMID:  20499188     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Nuclear transfer (NT) cloning involves manual positioning of individual donor-recipient cell couplets for electrofusion. This is time-consuming and introduces operator-dependent variation as a confounding parameter in cloning trials. In order to automate the NT procedure, we developed a micro-fluidic device that integrates automated cell positioning and electrofusion of isolated cell couplets. A simple two layer micro-fluidic device was fabricated. Thin film interdigitated titanium electrodes (300 nm thick, 250 microm wide and 250 microm apart) were deposited on a solid borosilicate glass substrate. They were coated with a film of electrically insulating photosensitive epoxy polymer (SU-8) of either 4 or 22 microm thickness. Circular holes ("micropits") measuring 10, 20, 30, 40 or 80 microm in diameter were fabricated above the electrodes. The device was immersed in hypo-osmolar fusion buffer and manually loaded with somatic donor cells and recipient oocytes. Dielectrophoresis (DEP) was used to attract cells towards the micropit and form couplets on the same side of the insulating film. Fusion pulses between 80 V and 120 V were applied to each couplet and fusion scored under a stereomicroscope. Automated couplet formation between oocytes and somatic cells was achieved using DEP. Bovine oocyte-oocyte, oocyte-follicular cells and oocyte-fibroblast couplets fused with up to 69% (n = 13), 50% (n = 30) and 78% (n = 9) efficiency, respectively. Fusion rates were comparable to parallel plate or film electrodes that are conventionally used for bovine NT. This demonstrates proof-of-principle that a micropit device is capable of both rapid cell positioning and fusion.
Authors:
Andrew L Clow; Paul T Gaynor; Björn J Oback
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Biomedical microdevices     Volume:  12     ISSN:  1572-8781     ISO Abbreviation:  Biomed Microdevices     Publication Date:  2010 Oct 
Date Detail:
Created Date:  2010-08-05     Completed Date:  2011-01-10     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  100887374     Medline TA:  Biomed Microdevices     Country:  United States    
Other Details:
Languages:  eng     Pagination:  777-86     Citation Subset:  IM    
Affiliation:
Department of Electrical and Computer Engineering, University of Canterbury, Private Bag 4800, Christchurch 8140, New Zealand. alc76@student.canterbury.ac.nz
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MeSH Terms
Descriptor/Qualifier:
Animals
Automation
Cattle
Cell Fusion / instrumentation*
Electrodes
Electrophoresis / instrumentation*
Female
Microfluidic Analytical Techniques*
Models, Theoretical
Nuclear Transfer Techniques / instrumentation*
Oocysts / cytology
Systems Integration

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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